We have identified and characterized two different sodium-coupled monocarboxylate cotransporters (SMCT) from zebrafish (Danio rerio), electrogenic (zSMCTe) and electroneutral (zSMCTn). zSMCTn is the 12th member of the zebrafish Slc5 gene family (zSlc5a12). Both zSMCT sequences have ∼50% homology to human SLC5A8 (hSMCT). Transport function and kinetics were measured in Xenopus oocytes injected with zSMCT cRNAs by measurement of intracellular Na+ concentration ([Na+]i) and membrane potential. Both zSMCTs oocytes increased [Na+]i with addition of monocarboxylates (MC) such as lactate, pyruvate, nicotinate, and butyrate. By using two electrode voltage clamp experiments, we measured currents elicited from zSMCTe after MC addition. MC-elicited currents from zSMCTe were similar to hSMCT currents. In contrast, we found no significant MC-elicited current in either zSMCTn or control oocytes. Kinetic data show that zSMCTe has a higher affinity for lactate, nicotinate, and pyruvate (Km L-lactate = 0.17 ± 0.02 mM, Kmnicotinate = 0.54 ± 0.12 mM at -150 mV) than zSMCTn (Km L-lactate = 1.81 ± 0.19 mM, Kmnicotinate = 23.68 ± 4.88 mM). In situ hybridization showed that 1-, 3-, and 5-day-old zebrafish embryos abundantly express both zSMCTs in the brain, eyes, intestine, and kidney. Within the kidney, zSMCTn mRNA is expressed in pronephric tubules, whereas zSMCTe mRNA is more distal in pronephric ducts. zSMCTn is expressed in exocrine pancreas, but zSMCTe is not. Roles for Na +-coupled monocarboxylate cotransporters have not been described for the brain or eye. In summary, zSMCTe is the zebrafish SLC5A8 ortholog, and zSMCTn is a novel, electroneutral SMCT (zSlc5a12). Slc5a12 in higher vertebrates is likely responsible for the electroneutral Na+/lactate cotransport reported in mammalian and amphibian kidneys.